The goal of this project is to fully characterize at the local and systemic levels T cell-mediated immune responses (CMI) induced by oral immunization with new generation attenuated S. typhi strains and to identify the cellular and molecular mechanisms that best correlate with protection against challenge with wild-type S. typhi. The development of improved typhoid vaccines has been hampered by a considerable lack of information on the specific determinants of protective CMI to S. typhi infection. Moreover, insufficient information is available regarding human mucosal immune responses to S. typhi, likely to be a key defense mechanism. Our central hypothesis is that the induction of potent and sustained CMI at both the local (e.g., the gut microenvironment) and systemic levels is critical for the development of an effective typhoid vaccine. Specifically, using peripheral blood mononuclear cells (PBMC) and mucosal biopsy specimens obtained from volunteers vaccinated with attenuated strains of S. typhi or challenged with wild-type S. typhi we propose. 1) To test the hypothesis that protective CMI responses against S. typhi will be determined by a set of immunodominant epitopes derived from S. typhi antigens. We will study the fine specificity of anti-S. typhi T cell responses by identifying immunodominant S. typhi proteins and T cell epitopes using a panel of T cell clones obtained from blood of immunized volunteers. 2) To test the hypothesis that CTL induced by immunization kills infected target cells by a combination of perforin and FAS ligand-mediated mechanisms. 3) To test the hypothesis that dendritic cells (DC) play a key role in the induction of protective immune responses in humans following oral immunization with attenuated strains of S. typhi. 4) To test the hypothesis that challenge with wild-type S. typhi or immunization with attenuated strains elicits the appearance in the gut mucosa of specific CTL effectors and T lymphocytes that produce Type-1 cytokines, and that these responses correlate with protection following challenge with wild-type S. typhi.